Cell migration is essential for a variety of physiological and pathological processes, such as angiogenesis, cancer metastasis, wound healing and inflammation. In the vascular system, significant efforts have focused on cell migration in the context of capillary morphogenesis. Through these studies, various mechanical and biochemical factors have been identified as critical in regulating endothelial cell migration and tube formation, such as chemotactic or chemokinetic effects of single or multiple growth factors1, interstitial fluid flow2,3 and matrix stiffness4,5. Despite the detailed understanding of individual components, how these factors are integrated to produce a specific cellular response has yet to be elucidated, creating the need for a versatile in vitro system in which these environmental factors can be studied in a controlled fashion. Achieving this will facilitate investigations that lead to a better understanding of how biochemical and mechanical factors act together in physiological and patho-physiological processes and ultimately contribute to improved tissue engineering and therapeutic strategies.
Understanding cell migration in capillary morphogenesis is therapeutically important because of its relation to human diseases and developmental phenomena6. Typical cell migration assays are unable to integrate complex environmental factors, particularly those that facilitate the formation of new tube-like structures from pre-formed capillaries or a cell monolayer within a three dimensional environment. One of the current capillary morphogenesis assays produces planar tubular networks on ECM-like substrates7-9. Capillary-like structures formed with this technique, however, have a reversed cell polarity with media on the outside and scaffolding materials on the inside7. Other approaches include sandwiching one cell monolayer between two layers of scaffold material8 and inducing capillary invasion by introducing chemical gradients9. These experiments have provided a foundation for understanding capillary morphogenesis, but are limited by an inability to image cell invasion in-plane, which would lead to more detailed characterization of the factors influencing this biological process.
Historically, many assays have been used to study cell migration10, such as the wound assay11,12, the TEFLON® fence assay13 and the Boyden chamber14,15. Both the wound assay and TEFLON® fence assay are limited to studying cell migration in 2D. The Boyden chamber mimics most closely the physiologic 3D environment, but is not conducive to quantifying cell migration in real time. Another assay with endothelial cell coated beads or spheroids embedded in collagen gel was able to generate tube-like structures in a three dimensional environment. The assay allowed the generation of stable tube-like structures and the co-culture with other cell types16-18, but the initial endothelial cell seeding surface is a rigid bead that does not allow for physiological factors such as a fluid-matrix interface and fluid flow experienced by endothelial cells in vivo. Furthermore, with the current assays, the chemokinetic and chemotactic effects are difficult to differentiate. In the context of cell migration, chemotaxis represents cells migrating towards the chemoattractant, while chemokinesis represents an increased motility in the presence of a particular biochemical factor. Due to technical difficulties in maintaining a controlled gradient, the two effects are not easily distinguished. Challenges to the existing techniques are: (i) to have precise control of the mechanical and biochemical factors in a physiologically-relevant condition, (ii) to have excellent optical resolution in real time, and (iii) to minimize sample variability and enhance sensitivity for quantification.
Microfabrication and microfluidic technology has the potential to overcome these challenges in studying cell migration by allowing for precise control of multiple environmental factors. However, current efforts in this area have continued to investigate isolated factors. For example, microfabricated patterns have enabled the demonstration of preferential migration in the direction of increased stiffness19,20. Microfluidic technology has also enabled the precise control of biochemical gradients and quantification of the resulting cell migrations21.